Novel signaling pathway for the production of inglammatory pain and neuropathy

ABSTRACT

This invention relates to a remedy for treating allergic contact dermatitis containing a substance having MEK inhibitory action as its effective component.

TECHNICAL FIELD

This invention relates to a remedy for treating allergic contactdermatitis.

BACKGROUND ART

Epidermal cells are well known to play an important roll in allergicdermatitis. More specifically, keratinocytes are activated by thestimuli of antigen and chemical substance to produce various cytokinessuch as granulocyte/macrophage colony-stimulating factor (GM-CSF) (Leeet al., J Immunol, 159: 5084-5088 (1997); and Steinhoff et al., CurrOpin Allergy Clin Immunol, 1: 469-476 (2001)). The thus producedcytokines activate Langerhans' cell which is an antigen presenting cell(Katz et al., Nature, 282: 324-326 (1979)), and the activatedLangerhans' cell moves to the regional lymph node where it presents theantigen to naive T lymphocyte (Kripke et al., J Immunol, 145: 2833-2838(1990); and Banchereau et al., Annu Rev Immunol, 18: 767-811 (2000)).

Increase in GM-CSF has been reported for the patients of atopicdermatitis (Pastore et al., J Clin Invest, 99: 3009-3017 (1997)), and ithas been conceived that GM-CSF from keratinocyte acts on Langerhans'cell to enhance its immunostimulative function (Witmer-Pack et al., JExp Med, 166: 1484-1498 (1987); Heufler et al., J Exp Med, 167: 700-705(1988); and Salgado et al., J Invest Dermatol, 113: 1021-1027 (1999))and simultaneously suppresses production of IL-12 which is a substancethat induces type 1 helper T cell involved in the contact dermatitis(Toda et al., J Immunol, 164: 5113-5119 (2000)) to induce immuneresponse by the type 2 helper T cell in the atopic dermatitis patients.Extracellular regulated kinase (ERK) which is one of themitogen-activated protein kinase (MAPK) is also known to be involved inthe production of GM-CSF (Kimata et al., Biochem Pharmacol, 60: 589-594(2000); Hallsworth et al., Am J Respir Crit Care Med, 164: 688-697(2001); and Dumitru et al., Cell, 103: 1071-1083 (2000)).

The signal transduction mediated by the MAPKs system takes specificpathways involving specific MAPK kinases, and exemplary signaltransduction pathway includes the route from MEK1/2 via ERK1/2, theroute from MKK7 and MKK4/SEK1 via JNK, and the route from MKK3/6 via p38(Yamanaka et al., Experimental medicine, 20:206-210 (2002)). With regardto the interaction of ERK and p38, MEK/ERK is believed to actcompetitively with p38 in view of the facts that DNA synthesis in ratlung myofibroblast by cell propagation stimulus and activation of ERK1/2are enhanced by SB203580 which is the selective inhibitor for p38 (Riceet al., Am J Respir Cell Mol. Biol, 27: 759-765 (2002)), and that, inthe formation of osteoclast, SB203580 and PD169316 which are specificinhibitors for the p38 exhibit suppressive action while U0126 andPD98059 which are specific inhibitors for the MEK exhibit promotiveaction, and that phosphorylation of the ERK is enhanced by the p38inhibitor, while phosphorylation of the p38 is enhanced by the MEKinhibitor (Hotokezaka et al., J Biol Chem, 277: 47366-47372 (2002)).

These findings indicate that the MEK/ERK-mediated pathway is involved inthe onset mechanism of the atopic dermatitis, while GM-CSF, whoseproduction is mediated by MEK/ERK, suppresses induction of Th1 which isinvolved in the allergic contact dermatitis. As described above, theonset mechanism of the atopic dermatitis is clearly different from thatof the contact dermatitis, and accordingly, development of thetherapeutic drugs targeting these two diseases had to be conducted froma different point of view.

DISCLOSURE OF THE INVENTION

While various drugs have been developed for atopic dermatitis, only feweffective agents are available for contact dermatitis. Accordingly, anobject of the present invention is to provide a therapeutic agent fortreating allergic contact dermatitis.

The inventors of the present invention made an extensive investigationin search of drugs effective in treating the allergic contactdermatitis, and quite unexpectedly found that an MEK inhibitor exhibitsexcellent therapeutic effects in mouse model of picryl chloride-induceddermatitis which is widely used as a model for allergic contactdermatitis, while MEK/ERK inhibitor has been conceived to be effectivefor atopic dermatitis but not for the contact dermatitis since action ofthe MEK/ERK is competitive to p38. The present invention has beencompleted on the basis of such a finding.

Accordingly, this invention provides a therapeutic agent for treatingallergic contact dermatitis containing a substance having MEK inhibitoryaction as its effective component.

This invention also provides use of a substance having MEK inhibitoryaction for production of a therapeutic agent for treating allergiccontact dermatitis.

This invention also provides a method for treating allergic contactdermatitis comprising administering a substance having MEK inhibitoryaction at an effective amount.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a view showing the therapeutic effects of U0126 for allergiccontact dermatitis. The results are represented as average ± standarderror. (*: p<0.05, N=8)

FIG. 2 is a view showing the therapeutic effects of PD98059 for allergiccontact dermatitis. The results are represented as average ± standarderror. (*: p<0.05, **: p<0.01)

BEST MODE FOR CARRYING OUT THE INVENTION

The effective component of the drug for treating allergic contactdermatitis of the present invention is not particularly limited as longas it is a substance having an inhibitory action for MEK, and exemplarysuch components include known MEK inhibitors such asbis[amino[(2-aminophenyl)thio]methylene]butanedinitrile (U0126, J. Biol.Chem., Vol. 273, No. 29, pages 18623-18632 (1998)) and2-(2-amino-3-methoxyphenyl)-4-oxo-4H-[1]benzopyran (PD98059, U.S. Pat.No. 5,525,625).

These MEK inhibitors may be used in the form of a salt, and the salt isnot particularly limited as long as it is a pharmaceutically acceptablesalt. Exemplary salts include acid adduct salts of a mineral acid suchas hydrochloride, hydrobromide, hydroiodide, sulphate, and phosphate;and acid adduct salts of an organic acid such as benzoate,methanesulphonate, ethanesulphonate, benzenesulphonate,p-toluenesulphonate, succinate, maleate, fumarate, tartarate, citrate,and acetate. Among these, the preferred are hydrochloride, sulphate,maleate, and fumarate.

These MEK inhibitors may also exist in the form of a solvate astypically represented by hydrate. Such solvate is also within the scopeof the invention.

These MEK inhibitors exhibit excellent therapeutic effects for allergiccontact dermatitis as will be demonstrated in the Examples, and they arequite useful as a drug for treating contact dermatitis such as skininflammation caused by immediate response as well as delayed responsewhich are induced by the contact with a chemical substance, nickel,rubber, or other minerals, animals, plants, fungi, and the like.

The drug for treating allergic contact dermatitis of the presentinvention contains a MEK inhibitor as its effective component,administration form is not particularly limited and any adequate formmay be selected depending on the object of the treatment. Exemplarydosage forms include oral preparation, injection, suppository, and also,percutaneous preparation, inhalant, eye drops, nasal drops, ear drops,and other external preparations, and the composition well suited forsuch administration form may be produced by any of the common methodsknown in the art by incorporating a pharmaceutically acceptable carrier.

When an oral solid preparation is prepared, the MEK inhibitor may beadmixed with an excipient, and optionally, with a binder, adisintegrant, a lubricant, a colorant, a taste corrective, a flavorcorrective, or the like, and the mixture may be produced into tablets,coated tablets, granules, powder, capsules, or the like by the methodcommonly used in the art. The additives used may be those commonly usedin the art, and exemplary excipients include lactose, sugar, sodiumchloride, glucose, starch, calcium carbonate, kaolin, microcrystallinecellulose, and silicic acid, and exemplary binders include water,ethanol, propanol, simple syrup, glucose solution, starch solution,gelatin solution, carboxymethylcellulose, hydroxypropyl cellulose,hydroxypropyl starch, methylcellulose, ethylcellulose, shellac, sodiumphosphate, and polyvinylpyrrolidone. Exemplary disintegrants include drystarch, sodium arginate, agar powder, sodium hydrogenccarbonate, calciumcarbonate, sodium laurylsulfate, monoglyceride stearate, and lactose,and exemplary lubricants include purified talk, stearate, borax, andpolyethyleneglycol. Exemplary colorants include β-carotene, yellowferric oxide, and caramel, and exemplary taste correctives include whitesugar, orange peel, citric acid, and tartaric acid. Exemplary flavorcorrective include diatomaceous earth, and kaolin.

When an oral liquid preparation is prepared, the MEK inhibitor may beadmixed with a taste corrective, a buffer, a stabilizer, a flavorcorrective, a preservative, or the like, and the mixture may be producedinto a liquid preparation, a syrup, a elixir, or the like by the methodcommonly used in the art. In this case, the taste and flavor correctivesmay be selected from those described above, and the buffer used may besodium citrate or the like. Exemplary stabilizers include gum traganth,gum arabic, and gelatin, and exemplary preservatives include methylparaoxybenzoate, ethyl paraoxybenzoate, and propyl paraoxybenzoate.

When an injection is prepared, the MEK inhibitor may be admixed with apH adjusting agent, a buffer, a stabilizer, an isotonic agent, a topicalanesthetic, or the like, and the mixture may be produced intosubcutaneous, intramuscular, or intravenous injection by the methodcommonly used in the art. In this case, the pH adjusting agent and thebuffer may be sodium citrate, sodium acetate, sodium phosphate, or thelike, and exemplary stabilizers include sodium pyrosulfite, EDTA,thioglycolic acid, and thiolactic acid. Exemplary isotonic agentsinclude sodium chloride and glucose, and exemplary topical anestheticsinclude procaine hydrochloride, and lidocaine hydrochloride.

When a suppository is prepared, the MEK inhibitor may be admixed with apharmaceutical carrier known in the art such as polyethyleneglycol,lanoline, cacao butter, or a fatty acid triglyceride, and if desired,further with a surfactant such as Tween™, and produced into the desiredpreparation by the method commonly used in the art.

When an external preparation such as ointment, cream, gel, lotion,solution, water-based poultice, or non-water-based poultice is produced,the MEK inhibitor may be admixed with a base, a water-soluble polymer, acrosslinking agent, a tackifier, a solvent, a stabilizer, a surfactant,a pH adjusting agent, a preservative, or the like commonly used with theMEK inhibitor, and the mixture may be stirred and produced into thedesired preparation by the method commonly used in the art. Exemplarybases include purified lanoline, liquid paraffin, white petrolatum,white Beeswax, octyldodecyl alcohol, and paraffin, and exemplarywater-soluble polymers include carboxyvinyl polymer, pullulan, sodiumcarboxy methyl cellose, sodium arginate, and xanthan gum. Exemplarycrosslinking agents include dihydroxyaluminum amino acetate, and driedaluminum hydroxide gel, and exemplary tackifiers include partiallyneutralized polyacrylic, poly(sodium acrylate), poly(2-ethylhexylacrylate), and styrene-isoprene-styrene block copolymer. Exemplarysolvents include glycerin, 1,3-butylene glycol, water, oleic acid,castor oil, and benzylalcohol, and exemplary surfactants include sodiumlauryl sulfate, glyceryl monostearate, sucrose fatty ester, andpolyoxyethylene fatty ester. Exemplary pH adjusting agents includesodium hydroxide, and citric acid, and exemplary preservatives includemethyl paraoxybenzoate, ethyl paraoxybenzoate, and propylparaoxybenzoate.

When eye drops are prepared, the MEK inhibitor may be admixed with a pHadjusting agent, a stabilizer, an isotonic agent, a preservative, or thelike, and produced into the desired preparation by the method commonlyused in the art. The additives used may be those commonly used in theart, and exemplary pH adjusting agent is sodium phosphate, and exemplarystabilizers include sodium pyrosulfite and EDTA. An exemplary isotonicagent is sodium chloride, and an exemplary preservative ischlorobutanol.

When nasal drops are prepared, the MEK inhibitor may be admixed with apH adjusting agent, a stabilizer, an isotonic agent, a preservative, orthe like, and produced into the desired preparation by the methodcommonly used in the art. The additives used may be those commonly usedin the art, and an exemplary pH adjusting agents is sodium phosphate,and exemplary stabilizers include sodium pyrosulfite and EDTA. Anexemplary isotonic agent is sodium chloride, and an exemplarypreservative is benzalkonium chloride.

When ear drops are prepared, the MEK inhibitor may be admixed with a pHadjusting agent, a buffer, a stabilizer, an isotonic agent, apreservative, or the like, and produced into the desired preparation bythe method commonly used in the art. The additives used may be thosecommonly used in the art, and exemplary pH adjusting agents and buffersinclude sodium phosphate, and exemplary stabilizers include sodiumpyrosulfite and EDTA. An exemplary isotonic agent is sodium chloride,and an exemplary isotonic agent is benzalkonium chloride.

The drug for treating allergic contact dermatitis of the presentinvention may be administered at a dose which may vary according to theage, body weight, symptoms, administration form, frequency of theadministration, and the like. However, the drug is generallyadministered orally or parenterally at 1 to 1000 mg as a MEK inhibitorper day per adult in one to several dosage(s).

EXAMPLES

Next, the present invention is described in further detail by referringto the Examples, which by no means limit the scope of the invention.

Example 1

A. Test method

(1) Sensitization and Induction of the Dermatitis

Female BALB/C mice purchased from Charles River Japan, Inc. were used.The mice were sensitized by spreading 0.1 mL of 7% picryl chloride (PC)solution in acetone on shaved abdomen, and 7 days after thesensitization, dermatitis was induced by spreading 0.01 mL of 1% PCsolution in acetone on the front and the back of the left auricle (i.e.0.02 mL in total).

(2) Measurement of the Dermatitis

Thickness of the auricle was measured before the dermatitis inductionand 24 hours after the dermatitis induction, and the difference in thethickness was measured.

(3) Preparation of the Test Substance and its Administration

The test substance used was U0126 which is known as a specific MEKinhibitor. U0126 was used by dissolving in dimethylsulfoxide. The testsubstance was applied on both front and back of the left auricle at anamount of 0.02 mL (0.04 mL in total) 1 hour before the dermatitisinduction.

(4) Data Processing

The results are represented as average ± standard error. Student's ttest was used for the determination of the significant difference, andsignificance level of less than 5% was considered significant.

B. Test Results

As shown in FIG. 1, U0126 (a specific inhibitor for MEK1/2 whichactivates extracellular regulated kinase (ERK1/2) which is a member ofMitogen-activated protein kinase (MAPKs)) exhibited suppressive actionfor allergic contact dermatitis in a dose dependent manner in the rangeof 0.1 to 10 μg/ear, and the action at 100 μg/ear was substantially thesame level as the action of 10 μg/ear.

Example 2

A. Test Method

(1) Sensitization and Induction of the Dermatitis

The procedure of Example 1 was repeated.

(2) Measurement of the Dermatitis

The procedure of Example 1 was repeated.

(3) Preparation of the Test Substance and its Administration

The procedure of Example 1 was repeated by replacing U0126 with PD98059.

B. Test Results

As shown in FIG. 2, PD98059 exhibited suppressive action for allergiccontact dermatitis.

1. A remedy for treating allergic contact dermatitis containing a substance having MEK inhibitory action as its effective component.
 2. A remedy according to claim 1 wherein the substance having MEK inhibitory action is bis[amino[(2-aminophenyl)thio]methylene]butanedinitrile or 2-(2-amino-3-methoxyphenyl)-4-oxo-4H-[1]benzopyran.
 3. A remedy according to claim 1 or 2 wherein the drug is administered by external administration.
 4. A remedy according to claim 1 or 2 wherein the drug is administered by percutaneous administration, inhalation, eye instillation, nasal instillation, or ear instillation.
 5. Use of a substance having MEK inhibitory action for production of a therapeutic agent for treating allergic contact dermatitis.
 6. Use according to claim 5 wherein the substance having MEK inhibitory action is bis[amino[(2-aminophenyl)thio]methylene]butanedinitrile or 2-(2-amino-3-methoxyphenyl)-4-oxo-4H-[1]benzopyran.
 7. Use according to claim 5 or 6 wherein the agent is administered by external administration.
 8. Use according to claim 5 or 6 wherein the agent is administered by percutaneous administration, inhalation, eye instillation, nasal instillation, or ear instillation.
 9. A method for treating allergic contact dermatitis comprising administering a substance having MEK inhibitory action at an effective amount.
 10. A method according to claim 9 wherein the substance having MEK inhibitory action is bis[amino[(2-aminophenyl)thio]methylene]butanedinitrile or 2-(2-amino-3-methoxyphenyl)-4-oxo-4H-[1]benzopyran.
 11. A method according to claim 9 or 10 wherein the administration is by external administration.
 12. A method according to claim 9 or 10 wherein the administration is by percutaneous administration, inhalation, eye instillation, nasal instillation, or ear instillation. 